The epigenome is the physiological template of the genetic information in all eukaryotic cells [Jenuwein, T. and Allis, C. D. (2001) Science 293:1074-1080]. The epigenome formats the genetic information by either presenting an accessible structure that can be expressed or by forming an inaccessible structure that cannot be accessed and thus is silenced. The epigenome comprises two major components: a group of proteins called histones that package the DNA into either accessible or inaccessible formats called chromatin, and a coating of methyl groups that are covalently bound to the DNA molecule itself. A long line of data has established that aberrations of the epigenome are a hallmark of all tumour cells, with one of the clear aberrations being global hypomethylation of DNA [Szyf, M. (1996) Pharmacol Ther 70:1-37].
DNA methylation is catalysed by the enzyme DNA methyltransferase and occurs via transfer of methyl groups from S-adenosyl methionine to the 5′ position of cytosine residues [Adams, R. L. et al. (1984) Curr Top Microbiol Immunol 108:142-156]. This process was previously believed to be irreversible, however, a protein exhibiting demethylase activity has recently been identified. The cDNA encoding this methylated DNA binding protein has been cloned and purified [Bhattacharya, S. K. et al. (1999) Nature 397:579-583], and the encoded DNA demethylase, also referred to as MBD2, has been shown to exhibit an activity that can reverse the DNA methylation reaction and hence act as a bona fide demethylase [Ramchandani, S. et al. (1999) Proc Natl Acad Sci USA 96:6107-6112; Scanlan, M. J. et al. (1998) Int J Cancer 76:652-658]. A possible role for DNA demethylase in cancer has been suggested. For example, a nonbiased screen for highly enriched antigens in humans with colorectal cancer uncovered a cDNA identical to that encoding the demethylase protein [Vilain, A. et al. (1999) FEBS Lett 460:231-234]. In addition, a recent study of demethylase mRNA expression in various breast cancer cell lines demonstrated a correlation between the level of genome hypomethylation, aberrant chromosomes and expression of the cDNA encoding demethylase [Hattori, M. et al. (2001) J Int Med Res 29:204-213]. Similarly, a correlation between malignancy, expression of MBD2/demethylase and demethylation in ovarian cancer has been demonstrated [Hattori, M. et al. (2001) Cancer Lett 169:155-164; Kanai, Y. et al. (1999) Biochem Biophys Res Commun 264:962-966].
Expression of a reverse-oriented full-length mbd2/dMTase cDNA in human embryonic kidney (HEK) cells has been shown to inhibit anchorage-independent growth of these cells in vitro [International Patent Application No. WO 99/24583]. However, such a large expression construct has limited use in vivo due to such factors as rapid degradation of the expressed transcript by nucleases and poor bioavailability.
This background information is provided for the purpose of making known information believed by the applicant to be of possible relevance to the present invention. No admission is necessarily intended, nor should be construed, that any of the preceding information constitutes prior art against the present invention.